Effect of fasting on uteroplacental amino acid metabolism in the pregnant sheep.

In the fed state, the net utilization of most amino acids by the ovine uteroplacental unit (defined as uterine uptake minus umbilical uptake) is zero. This study was to determine if this is true for the fasted pregnant ewe also, when uteroplacental glucose uptake is diminished. Glucose and amino acid concentrations were determined in 9 pregnant ewes across the uterine and umbilical circulations. Uterine and umbilical blood flows were determined by the steady state antipyrine diffusion method, and net uptakes across each circulation calculated by application of the Fick principle. Five amino acids were found to have a consistently greater uterine uptake than umbilical uptake. In addition, glutamate had a negative net umbilical uptake coupled with no net uterine uptake. Fasting resulted in an increase in the differences between the respective uptakes for all five amino acids. No additional amino acids were found to have disparate uptakes than were seen in the fed state. Umbilical glutamate uptake became less negative during fasting, but the difference between the uterine and umbilical circulations remained unchanged, as the uterine circulation took up glutamate during fasting. The umbilical circulation took up alpha-ketoisocaproate 0.626 +/- 0.18, fed), and the uptake was increased during fasting (1.8 +/- 0.6). These findings suggest that the branched chain amino acids and glutamine are utilized by the ovine uteroplacenta to a greater degree than are most other amino acids, and at increased rates during fasting of the ewe. These amino acids, through deamination, may be the source of uteroplacental ammonia production, as well as serving as an energy substrate when glucose supply is limited.     

Gluconeogenesis from lactate in the chronically catheterized baboon fetus

Gluconeogenesis from lactate may be qualitatively identified in the chronically catheterized baboon fetus in the maternal fed and fasted state. Infusion of 250 microCi U-14C-lactate to the fetus over a 150-min period leads to the appearance of 14C-glucose in the fetal circulation. Little 14C-lactate or glucose appears in the maternal circulation, supporting fetal production of glucose from lactate. Maternal glucose infusion seems to inhibit fetal gluconeogenesis. The mean plateau in percent disintegrations per minute of glucose compared to lactate in the maternal fed state is 14.7 +/- 2.2 compared to 16.1 +/- 3.0 in the maternal fasting state and 11.1 +/- 0.6 during maternal glucose infusion. It is clear that the primate fetus is capable of gluconeogenesis before term. Quantitation of this capacity awaits development of a model permitting assessment of maternal-placental and fetal substrate flux.     

Diurnal changes in plasma amino acids in maple syrup urine disease

Protein turnover is a cyclic process with a net loss of protein in the (catabolic) fasted state and a net gain in the (anabolic) fed state. In maple syrup urine disease (MSUD) the early block of degradation of the branched-chain amino acids (BCAA) brings about the opportunity for evaluation of the diurnal variation in net protein anabolism and catabolism by studying cyclic changes in the plasma concentrations of BCAA. The alterations in plasma BCAA in a 3-y-old boy with classical MSUD were studied in the fed and fasted state over a period of 19 months. For each amino acid a total of 34 data pairs was calculated. The plasma concentrations of the BCAA leucine, valine and isoleucine were constantly higher in the fasted than in the fed state. Plasma concentrations of alloisoleucine, being a non-protein amino acid, did not participate in cyclic changes. In contrast, the essential amino acid pair tyrosine and phenylalanine increased after meals. The fasting concentration of alanine increased after feeding, while glycine did not change significantly. Healthy subjects show a decrease in all amino acids in the fasted (mild catabolic) state and an increase in the fed state. These findings in MSUD suggest a net decrease in non-BCAA as result of a greater rate of amino acid oxidation rate than of protein breakdown and a net entry of BCAA into plasma in the fasted state due to the specific metabolic block. Such changes in amino acid plasma pools have to be taken into account during monitoring of treatment and especially when in vivo leucine oxidation is assessed.     

Ovine fetal and maternal glycogen during fasting.

The present study was undertaken to assess the role of hepatic glycogen metabolism in fetal and maternal glucose homeostasis during a prolonged fast in the pregnant ewe. A control fed group of 13 ewes and 16 fetuses were compared to a 5-day-fasted group of 13 ewes and 17 fetuses, studied at 125 days gestation (term = 147 days). Tissue samples were obtained during pentobarbital anesthesia and frozen in liquid nitrogen. Protein, glycogen, active phosphorylase and total phosphorylase activity were determined. Fetal weight (3.61 vs. 2.86 kg) was decreased in the fasted group (p less than 0.001) while fetal hepatic glycogen was unchanged (59.8 vs. 52.4 mg/g tissue). Maternal liver glycogen decreased during fasting (38.2 vs. 4.0 mg/g tissue, p less than 0.001). Fetal active phosphorylase and total phosphorylase did not change between fed and fasted states (fed active phosphorylase 398 vs. fasted 441 and fed total phosphorylase 510 vs. fasted 574 mumol/h/g tissue). The maternal active phosphorylase and total phosphorylase decreased between fed and fasted (active phosphorylase 690 vs. 238 and total phosphorylase 981 vs. 599 mumol/h/g tissue, p less than 0.001). During fasting, the pregnant ewe depletes her hepatic glycogen stores, associated with a reduction in glycogen catabolizing enzyme activity. The fetus maintains a relatively large glycogen catabolizing enzyme activity, a relatively large glycogen reserve and substantial phosphorylase activity.     

Fetal and maternal amino acid concentrations during fasting in the ewe

The present investigation was undertaken to quantitate the whole-blood amino acid concentrations of the mother and fetus over an extended period of fasting using a chronic sheep preparation. Fetal amino acid levels generally increased or remained relatively high during the fast (e.g., alanine, glycine, valine, leucine). In contrast to the situation in the fetus, most amino acids (alanine, glycine, glutamine, serine, and threonine) decreased in concentration in the ewe with fasting. Alanine levels changed most dramatically in the ewe, decreasing to less than 50% of the fed state values within 72 h of fasting. Therefore, while the supply of gluconeogenic amino acids becomes limited within the maternal circulation during fasting, levels of these amino acids remain high or increase in the fetal arterial blood.     

Branched-chain amino acid carbon and nitrogen arteriovenous concentration differences across the ovine fetal hindlimb

During fasting, branched-chain amino acids (BCAA) are thought to be major sources of nitrogen for myocyte synthesis of alanine (Ala) and glutamine (Gln), as well as possible sources of carbon skeleton for Gln synthesis. To study the relationships between Ala, Gln, and BCAA, we utilized the chronic fetal lamb preparation and measured arteriovenous concentration differences of Ala, Gln, BCAA, and branched-chain alpha-keto acids across the fetal hindlimb. Studies were performed when the ewe was fed and repeated after 1 and 5 days of complete maternal fasting. Ala and Gln are released from fetal hindquarters during fasting (arteriovenous -9.6 +/- 5 and -8.8 +/- 4.1 mumol/liter), while arteriovenous differences for BCAA simultaneously increase by 65% as compared to the fed state. During fasting, total nitrogen exiting fetal hindlimb as Ala and Gln equals nitrogen entering as BCAA. Branched-chain alpha-keto acids are released from fetal hindquarters during the fed state as well as after 1 day of fasting; at 5 days of fasting only keto-isovalerate had a net negative arteriovenous difference. In all cases, the release was much smaller than the entry of the corresponding amino acid, as determined by simultaneously measured arteriovenous concentration differences. These results indicate: Ala and Gln are released from fetal hindlimb, as in postnatal animals, during fasting, shuttling nitrogen and carbon to liver and/or other tissues. There is negligible movement of BCAA carbon as the branched-chain alpha-keto acids from hindlimb to other tissues in the fetus. BCAA carbon taken up is utilized within the hindquarters. It may be used for tissue synthesis, as an energy source, or possibly for Gln synthesis.     

Cerebral blood flow and metabolism in fasting neonatal piglets

Cerebral blood flow and cerebral metabolism were studied in 8 fasting neonatal piglets. Cerebral blood flow was not significantly different (p greater than 0.1) at 24 h (75.0 +/- 30.7 ml/min/100 g) and 48 h (74.2 +/- 32.3 ml/min/100 g) of age. Blood glucose concentration was lower in 48-hour-old fasted piglets (1.27 +/- 0.60 mmol/l) in comparison with 24-hour-old fasted animals (2.90 +/- 0.52 mmol/l), and cerebral glucose utilisation rate was also reduced in the 48-hour-old piglets (17.6 +/- 6.0 mumol/min/100 g at 48 h, 34.2 +/- 12.2 mumol/min/100 g at 24 h). Blood lactate concentration and cerebral lactate utilisation rate were not significantly different at 24 and 48 h of age. Brain lactate utilisation accounted for approximately 10% of cerebral oxygen utilisation at 48 h in fasted piglets. Cerebral glucose and lactate utilisation at 48 h accounted for approximately 66% of cerebral oxygen uptake, indicating that other substrates make a major contribution to cerebral metabolic requirements in fasting neonatal piglets.     

Glucose utilization by the placenta and fetal tissues in fed and fasted pregnant rabbits

Glucose utilization by the placenta and individual fetal tissues was studied in vivo in conscious pregnant rabbits at 29 days of gestation. In the fed state, the rate of glucose utilization was similar in the placenta and the gravid uterus, suggesting that the rate of fetal glucose utilization was approximately 40 nmol/min/g. A 96-h maternal fast induced a significant decrease in glucose utilization by the myoendometrium and in the glucose utilization index by fetal liver and brown adipose tissue. No modification was observed in other fetal tissues. These results indicate that glucose utilization by the placenta and the whole fetus from 96-h fasted rabbits does not decrease despite profound changes in endocrine and metabolic maternal parameters.     

Amino acid metabolism in the human fetus at term: leucine, valine, and methionine kinetics

Human fetal metabolism is largely unexplored. Understanding how a healthy fetus achieves its fast growth rates could eventually play a pivotal role in improving future nutritional strategies for premature infants. To quantify specific fetal amino acid kinetics, eight healthy pregnant women received before elective cesarean section at term, continuous stable isotope infusions of the essential amino acids [1-13C,15N]leucine, [U-13C5]valine, and [1-13C]methionine. Umbilical blood was collected after birth and analyzed for enrichments and concentrations using mass spectrometry techniques. Fetuses showed considerable leucine, valine, and methionine uptake and high turnover rates. α-Ketoisocaproate, but not α-ketoisovalerate (the leucine and valine ketoacids, respectively), was transported at net rate from the fetus to the placenta. Especially, leucine and valine data suggested high oxidation rates, up to half of net uptake. This was supported by relatively low α-ketoisocaproate reamination rates to leucine. Our data suggest high protein breakdown and synthesis rates, comparable with, or even slightly higher than in premature infants. The relatively large uptakes of total leucine and valine carbon also suggest high fetal oxidation rates of these essential branched chain amino acids.     

Effect on the fetus of infusing a commercial amino acid preparation into a pregnant sheep

The common clinical practice of intravenous feeding of the pregnant woman poses the question of the effect on the fetus of such infusions. We have used the sheep as a model to study the change in fetal amino acid levels after a maternal infusion of Synthamin 13. The maternal plasma aminogram largely reflects the amino acid pattern in the infusate. However, in the fetal circulation only the branched chain amino acids (leucine, isoleucine and valine), phenylalanine and alanine rose significantly after infusion. Only leucine and isoleucine were observed to spill into the fetal urine. The results suggest that the ovine placenta selectively modifies the amino acid profile presented to the fetus when the maternal plasma aminogram is distorted. However, the fetus is not totally protected from changes in phenylalanine, which in high concentrations, is detrimental to normal development.     

Leucine kinetics during feeding in normal newborns

To examine how leucine and protein metabolism is affected by feeding, leucine kinetics were determined in 11 normal term newborns during feeding using a prime constant tracer infusion of 1-13C leucine combined with respiratory calorimetry. Fed newborns were compared with previously studied fasting newborns. Feeding and fasting newborns had similar rates of leucine oxidation (34 +/- 3 mumol/kg/h versus 31 +/- 4 mumol/kg/h) and leucine release from existing protein (156 +/- 16 mumol/kg/h versus 164 +/- 8 mumol/kg/h). In contrast, nonoxidative disposal rates of leucine (a reflection of protein synthesis) were significantly greater in feeding newborns (170 +/- 13 mumol/kg/h versus 129 +/- 9 mumol/kg/h). A significant positive correlation between birth weight and leucine flux was demonstrated in both feeding and fasting newborns. These results suggest that 1) newborns may accomplish protein accretion primarily by increases in protein synthesis rather than suppression of protein breakdown; 2) an estimate can be made of the minimal leucine intake required to replace irreversible leucine oxidative losses (816 mumol/kg/d, 107 mg/kg/d); and 3) the positive correlation between birth weight and leucine flux in both feeding and fasting newborns may be a result of differences in previous protein and energy supplies.     

Effects of fasting on gluconeogenic enzymes in the ovine fetus

Fetal and maternal sheep were studied to determine whether changes in gluconeogenic enzyme activities could be detected in the liver and/or kidney associated with maternal nutritional deprivation. Thirteen ewes and 16 fetuses were sacrificed in the fed state, while 13 ewes with 17 fetuses were sacrificed after 5 days of fasting, all at 125 days gestation (term = 147 days). Fetal weight was decreased in the fasted versus fed group (2.86 +/- 0.56 versus 3.61 +/- 0.58 kg, p less than 0.001). Tissues were analyzed for glucose-6-phosphatase, fructose-1,6-diphosphatase, pyruvate carboxylase, phosphoenolpyruvate carboxykinase, glutamate oxaloacetate aminotransferase, and glutamate pyruvate aminotransferase. In maternal liver, four of the six enzymes increased significantly during fasting, whereas none of the enzymes increased in maternal kidney. In fetal hepatic tissue, five of the six enzymes (with the exception of pyruvate carboxylase) increased during maternal fasting and three of the enzymes increased in renal tissue. These data are consistent with the potential for increased rates of gluconeogenesis in the ovine fetus during periods of compromised maternal nutrition.     

Effects of fasting on glucose turnover rate and metabolite levels in conscious pregnant guinea pigs

The effect of fasting during pregnancy is of particular interest in the guinea pig because of the large fetal mass carried to term. The present studies examined the effect of acute and chronic starvation on maternal glucose turnover in the guinea pig. In the first experiment, 7 near-term pregnant guinea pigs were fasted for 6 h. The maternal glucose concentration and glucose production decreased rapidly, falling to about 65-70% of fed levels at 4 h of starvation. Mothers demonstrated a 2.6-fold elevation in ketoacids after 2, 4 and 6 h starvation. In a second experiment, 5 non-pregnant and 11 near-term pregnant animals were studied in the control period and after 24 h of fasting. The maternal glucose concentration in the control state was independent of fetal mass. The maternal glucose turnover rate in the fed state correlated linearly with fetal mass. After 24 h of fasting, the glucose concentration and glucose turnover rate both decreased, with the magnitude of each decrease proportional to fetal mass. We conclude that, in the pregnant guinea pig, the fetal mass impacts significantly on maternal glucose metabolism in the fed and fasting states.     

The effect of euglycemic hyperinsulinemia on cerebral cortical glucose metabolism in newborn beagles

There is a paucity of information on the significance of insulin on neonatal cerebral glucose metabolism. The effect of insulin on neonatal cerebral glucose uptake and cerebral cortical metabolic intermediates was investigated with the euglycemic hyperinsulinemic clamp in unanesthetized beagles during the first day of life. Insulin was infused at various rates to sustain an elevated steady state plasma insulin concentration in individual pups. Furthermore, blood glucose and 2-deoxyglucose levels were also maintained ("clamped") in a steady state by infusion of glucose and 2-deoxy-[14C]-glucose. Mean (+/- SD) plasma insulin levels were 20 +/- 12 and 2971 +/- 3386 (33-14330) microU/ml in control and hyperinsulinemic pups. Blood glucose concentration was 4.43 +/- 2.64 mM during basal periods and 4.54 +/- 2.87 mM during the clamp period in study pups. Basal fasting glucose utilization in study pups was 43.9 +/- 24 mumol/kg/min and increased to 60.9 +/- 35.2 mumol/kg/min (p less than 0.001) during hyperinsulinemia. Immediately after the euglycemic hyperinsulinemic clamp or fasting in control pups, the cerebral cortex was frozen to the temperature of liquid nitrogen. No differences were noted for any cerebral cortical intermediate between the two pup groups. In addition, there was no relationship between the cerebral intermediates concentration when analyzed as a function of plasma insulin levels. The uptake of cerebral 2-deoxyglucose was analyzed as a function of plasma insulin concentration (120-6900 microU/ml). Brain tissue demonstrated a positive linear relationship for 2-deoxyglucose uptake as a function of plasma insulin concentration.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in 4E-BP1 and p70S6K phosphorylation in skeletal muscle of the ovine fetus after prolonged maternal fasting: effects of insulin and IGF-I

This study was conducted to investigate fasting-induced alterations in insulin signaling to the regulatory components of the translation machinery. Insulin (890 mIU/h) and IGF-I (40 nM/h) were infused into a chronically catheterized ovine fetus (0.85 gestation) for 7 h following a 5-d maternal fast. Amino acid and glucose concentrations were clamped to minimize the effects of alterations in circulating substrate concentrations. The IGF-I induced increase in 4E-BP1 phosphorylation (percentage in the gamma form) increased from 28% in control to 44% (NS). The insulin-induced increase in 4E-BP1 phosphorylation was more pronounced, and the gamma percentage was 56% on average in the insulin group. The insulin-induced increase in 4E-BP1 phosphorylation was lower than in fed animals and did not result in significant changes in eIF4E.4E-BP1 binding or eIF4E.eIF4G binding. Insulin increased PKB/Akt phosphorylation and p70S6K phosphorylation to a similar extent as in fed animals. We conclude that maternal fasting resulted in reduced insulin sensitivity of 4E-BP1 phosphorylation and eIF4F formation. This reduced insulin-induced 4E-BP1 phosphorylation was not due to a global defect in insulin signaling; the defects underlying the reduced basal phosphorylation and insulin-responsiveness of 4E-BP1 in fasted animals may be in signaling components other than, or downstream of, PKB/Akt. Selective inhibition of downstream components of insulin signaling allows fetuses to adapt to nutritional stress by decreasing the anabolic response to insulin and other growth factors, so that more amino acids can be used as oxidative substrate to compensate for shortage of energy due to reduced glucose supply.     

Effects of early amino acid administration on leucine and glucose kinetics in premature infants

We previously showed that, in prematurely born infants, an anabolic state without metabolic acidosis can be achieved upon intravenous amino acid (AA) administration in the immediate postnatal phase, despite a low energy intake. We hypothesized that the anabolic state resulted from an increased protein synthesis and not a decreased proteolysis. Furthermore, we hypothesized that the energy needed for the higher protein synthesis rate would be derived from an increased glucose oxidation. To test our hypotheses, 32 ventilated premature infants (<1500 g) received intravenously either solely glucose or glucose and 2.4 g AA/kg/d immediately postnatally. On postnatal d 2, each group received primed continuous infusions of either [1-13C]leucine or [U-13C6]glucose. 13CO2 enrichments in expiratory air and plasma [1-13C]alpha-KICA (as an intracellular leucine precursor) and [U-13C6]glucose enrichments were measured by mass spectrometry techniques. The AA administration resulted in an increased incorporation of leucine into body protein and a higher leucine oxidation rate, whereas leucine release from proteolysis was not affected. Glucose oxidation rate did not increase upon AA administration. In conclusion, the anabolic state resulting from AA administration in the immediate postnatal period resulted from increased protein synthesis and not decreased proteolysis. The energy needed for the additional protein synthesis was not derived from an increased glucose oxidation.     

Effect of uterine artery insulin infusions on umbilical glucose uptake in sheep

Although glucose is an important fuel for fetal oxidative metabolism, regulation of its availability to the mammalian fetus is poorly understood. This study was performed to determine the effect of infusions of insulin into the uterine arterial circulation on umbilical uptake of glucose in chronically instrumented, unstressed sheep. Twenty-eight determinations of umbilical glucose uptake and diffusion clearance of glucose by the placenta were made in four ewes. Immediately following a control study during which saline was infused into the uterine artery, porcine regular insulin diluted in saline was infused at 0.05 to 8.1 mU/min . kg uterine weight for 20--30 min and the determinations were repeated. Subsequent studies were performed at the conclusion of additional infusions of insulin to a maximum of 21.6 mU/min . kg. There was a significant increase in umbilical glucose uptake during initial insulin infusions (4.47 +/- 0.6 mg/min . kg fetus) compared to the control studies (3.08 +/- 0.6 mg/min . kg) associated with an increase in diffusion clearance (13.8 +/- 1.9 ml/min . kg fetus vs. 8.99 +/- 1.8 ml/min . kg). When the total cumulative dose of exogenous insulin, It, was 162 mU/kg uterine weight or less, the umbilical uptake of glucose, Q, may be expressed as a function of maternal arterial blood glucose concentration in milligrams per dl, [A], and of It.     

Effect of maternal fasting on ovine fetal and maternal branched-chain amino acid transaminase activities

Activities of branched-chain amino acid transaminase were assayed in maternal skeletal muscle, liver and fetal skeletal muscle, cardiac muscle, liver, kidney and placenta obtained from fed and 5-day-fasted late gestation ewes. Very high activities were found in placenta; fetal skeletal muscle also had high activity. Fetal brain had intermediate activity, followed by cardiac muscle and kidney. Fetal liver possessed negligible activity. Activities were low in both maternal liver and skeletal muscle. Trends were seen for fasting to increase activities in fetal placenta, skeletal muscle, brain, kidney, heart and maternal liver, but these changes were statistically significant only for fetal brain and placental tissue. Fetal skeletal muscle activity was 100 times that of maternal skeletal muscle. These data imply differences in the metabolism of the branched-chain amino acids by fetal and adult ruminants and expand the thesis that branched-chain amino acids are important to the metabolism of the ovine fetus.     

Bidirectional placental transfer of glucose and its turnover in fetal and maternal sheep.

Glucose biokinetics were assessed simultaneously in the pregnant ewe and its fetus by a primed constant infusion of 2-3H glucose and U-14C glucose. Late in gestation fetal glucose turnover was 27.3 +/- 3.7 mg/min; expressed in terms of fetal weight this is 6 to 10 mg/kg/min. In the fed state the results indicated that all of the fetal glucose turnover was derived from the mother via placental transfer and there was no evidence that the fetus was capable of glucose production. Maternal glucose turnover was 145.6 +/- 9.3 mg/min (2.8 mg/kg/min). There was a significant amount of glucose (16.3 +/- 2.3 mg/min) transferred from the fetus to the mother. This feto-maternal transfer of glucose accounted for 11% of the maternal glucose turnover and approximately 50% of the total glucose coming to the fetus from the mother. This study provides the first in vivo simultaneous quantification of the bidirectional glucose transfer across the placenta.     

Amino acid interconversions in the fetal-placental unit: the animal model and human studies in vivo

Fetal growth and development are dependent upon the adequate provision of oxygen and substrates from the maternal circulation. The need for amino acids is related to protein synthesis, interconversion to other substrates, and oxidation. Amino acids cross the placenta by active transport systems, and their concentrations in the fetus are higher than in the mother. In addition, most amino acids are extensively metabolized within the placenta, and, for some nonessential amino acids, placental synthesis has been demonstrated in chronically catheterized fetal lambs. Interorgan cycling between the fetal liver and placenta has been hypothesized for nonessential amino acids like glycine and serine. Amino acids are oxidized within the fetal tissues, particularly in liver and muscle, with differences between amino acids and in relation to metabolic state. In human pregnancies, maternal-fetal transfer rates have been investigated in vivo by stable isotope methodologies performed at fetal blood sampling. The transfer rate of nonessential amino acids like glycine is significantly lower than for essential amino acids like leucine, confirming glycine synthesis in the fetoplacental unit also in human pregnancies. Moreover, when a steady state model is applied, the fetal-maternal ratio for [1-(13)C]leucine is significantly reduced in pregnancies associated with intrauterine growth restriction, reflecting a decrease in leucine placental transfer and/or an increase in protein catabolism in the fetoplacental unit. This reduction is proportional to the degree of severity of intrauterine growth restriction but is significant also in those intrauterine growth-restricted fetuses with normal oxygenation and acid-base status.